Szakács Zoltán, Noszál Béla
Department of Inorganic and Analytical Chemistry, Institute of Chemistry, Eötvös Loránd University, Budapest, Hungary.
Electrophoresis. 2006 Sep;27(17):3399-409. doi: 10.1002/elps.200600128.
Pressure-assisted CE (PACE) was applied to determine the previously inaccessible complete set of pK values for folic acid and eight related multiprotic compounds. PACE allowed the determination of all acidity macroconstants at low (<or=0.1 mM) concentration without interferences of selfassociation or photodegradation throughout the pH range. The accuracy of the constants was verified by NMR-pH, UV-pH, and potentiometric titrations and the data could be converted into physiological ionic strength. It was shown that even three overlapping pK values can be determined by CE with good precision (<0.06) and accuracy if an appropriately low sample throughput is used. Experimental aspects of PACE for the quantitation of acid-base properties are analyzed. The site-specific basicity data obtained for folic acid and methotrexate (MTX) reveal that apparently slight constitutional differences between folic acid and MTX carry highly different proton-binding propensities at analogous moieties, especially at the pteridine N(1) locus, providing straightforward explanation for the distinctive binding to dihydrofolate reductase at the molecular level.
压力辅助毛细管电泳（PACE）被用于测定叶酸及八种相关多质子化合物之前无法获得的完整pK值集。PACE能够在低浓度（≤0.1 mM）下测定所有酸度宏观常数，且在整个pH范围内不受自缔合或光降解的干扰。通过核磁共振 - pH、紫外 - pH和电位滴定法验证了常数的准确性，并且数据可以转换为生理离子强度。结果表明，如果使用适当低的样品通量，即使是三个重叠的pK值也能通过毛细管电泳以良好的精密度（<0.06）和准确度进行测定。分析了用于酸碱性质定量的PACE的实验方面。叶酸和甲氨蝶呤（MTX）获得的位点特异性碱度数据表明，叶酸和MTX之间明显细微的结构差异在类似部分具有高度不同的质子结合倾向。特别是在蝶啶N(1)位点，这在分子水平上为与二氢叶酸还原酶的独特结合提供了直接解释。
